ISSR和SRAP标记技术在葫芦科植物种质资源研究中的应用

阐述了简单重复序列间扩增(ISSR)和相关序列扩增多态性(SRAP)分子标记技术的基本原理和特点,综述与探讨了分子标记技术在葫芦科植物种质资源的亲缘关系鉴定与遗传多样性分析、分子遗传图谱的构建与目的性状基因的标记定位、基因库的构建和基因克隆、辅助育种选择及品种纯度鉴定等研究领域的广泛利用与推广,以期更合理高效地保护和利用葫芦科植物种质资源。     

用基于PCR的分子标记分析甘蓝型油菜抗感菌核病品系间的遗传距离

用离体叶接种法鉴定了甘蓝型油菜选系对菌核病的抗感性，结果表明绝大多数抗性品系与感病品种间的差异达到显著或极显著水平。以两种基于ＰＣＲ的标记估计所选１０ 份材料间的遗传距离。１９ 个随机引物扩增出了２６ 条重复性很好的ＲＡＰＤ 标记，１ 对依据植物Ｒ基因的ＮＢＳ区段设计的简并引物扩增出了３ 条多态性的带。依据２９ 个标记进行聚类分析，结果表明抗、感性材料在遗传距离为０．１２０６ 处被明显地区分开。５份抗性材料中，宁ＲＳ—１ 与中油８２１、中Ｒ８８８ 等其它抗性品系的遗传距离较远     

利用SRAP分子标记技术鉴定大白菜种子纯度

以大白菜品种多抗3及其父母本为研究材料，利用SRAP（相关序列扩增多态性）分子标记方法，通过对54对SRAP标准引物的筛选，获得了能区分大白菜多抗3及其亲本的引物Me4F/Em3R。采Me4F/Em3R引物对对5份多抗3系列商品种子纯度进行检验，种子的纯度分别为75.5%、72%、80.5%、68%和73%，与田间种植纯度鉴定结果符合率分别为94.6%、89.3%、93.2%、95.5%和91.25%。说明利用SRAP分子标记方法对多抗3大白菜一代杂种进行快速、准确的纯度鉴定是可行的。     

快速大量提取甜菜DNA的新方法

以甜菜幼嫩叶片为材料,通过冷冻真空干燥将材料变成干粉,利用CTAB法对DNA提取程序进行了研究,建立了一种简单、快速的甜菜DNA提取程序,既节省时间,又可获得高质量的DNA,可以满足SSR及SRAP扩增的需要。     

Occurrence of microsatellites in spinach sequences from computer databases and development of polymorphic SSR markers

Microsatellites are valuable tools as molecular markers in plant breeding. To establish genetic linkage maps or for population studies, information about the occurrence and usability of microsatellite markers in different species is necessary. Sequences of spinach Spinacia oleracea from computer databases were therefore searched for the presence of microsatellites. Sixty simple sequence repeats were found in 237 spinach sequences with a total of 349.4 kb DNA. After removing duplicated sequences, 50 different microsatellites with various motifs remained. Differences between nuclear and chloroplast DNA were not in the number of microsatellites but in their type and length. Chloroplast sequences from spinach contain only short strings of A and AT repeats, whereas nuclear sequences show a wider variety of motifs. Flanking primers for polymerase chain reaction (PCR) analysis were designed for 13 of these microsatellites and tested with two different varieties of spinach. Twelve primer pairs gave amplification products and seven of these showed polymorphisms in the variety ‘Wiremona’ but only one in the variety ‘Monatol’. These markers may be used for linkage analysis or population studies in spinach.     

Development of Sequence-characterized DNA Markers Linked to Temperature Insensitivity for Fruit Production in Longan (Dimocarpus longan Lour.) Cultivars

In this paper, we characterize a temperature dependence in longan (Dimocarpus longan Lour.) cultivars using the high annealing temperature random‐amplified polymorphic DNA (HAT‐RAPD) methodology. This form of genomic analysis allows a random sampling of the complete genome to provide information about phenotypic traits of interest. Using a set of 18 unique decamer primers, polymorphic bands ranging from 100 to 2500 bp were examined for 14 longan varieties from which banding patterns of interest were selected for conversion to the more reproducible and robust sequence characterized amplified region (SCAR) markers. In particular, one SCAR marker produced an electrophoresis banding pattern which could distinguish between longan varieties requiring a sustained interval at low temperatures for fruit production vs. those that do not. This band was composed of a nucleotide region 275 bases in length (DQ539047) in which the initial 90 nucleotides were >85 % identical to the 12‐oxophytodienoic acid (OPDA) reductase gene. OPDA is an immediate precursor to jasmonic acid which has been described as the ‘master switch’ for lipid‐derived environmental signalling to factors such as flowering and osmotic stress. The possible involvement of the jamonic acid signalling pathway could explain the recent success of potassium chlorate to induce off‐season flowering and fruit formation.     

广灵驴分子系谱的建立及群体遗传结构分析

旨在对广灵县优种驴场保种群体进行调查的基础上构建分子系谱,并对其种群的遗传结构进行分析。本研究采集保种群成年、体况良好的广灵驴（体重350~400 kg）颈静脉血10 mL（n=107）,其中公驴13份,母驴94份,抗凝处理后提取全血DNA。采用12个微卫星标记进行荧光PCR扩增后,用ABI3730测序仪进行分型。分型结果采用Cervus 2.0和Pedigraph 2.4软件构建分子系谱,同时采用STRUCTURE2.3和Fstat软件计算群体遗传参数,采用R语言的hclust函数绘制7头公驴及其后代的系统发生NJ树（邻接树）。结果,对107头种驴进行了分子系谱构建,找到了30头子代的父亲和7头子代的母亲,系谱可靠性>90%;微卫星标记的平均观测杂合度（H_Obs）和多态信息含量（PIC）分别为0.676 5和0.593 9,标记遗传多样性较高;NJ树对7个公驴家系进行了聚类;群体分化系数（F_ST）为0.184,群体平均近交系数（F_IT）为0.033,群体内近交系数（F_IS）为-0.238,且群体处于哈代-温伯格平衡状态,存在很弱的近交。本研究建立了广灵驴保种群可靠性较高的分子系谱并对其遗传结构进行了分析,证明该群体遗传多态性较高,群体近交系数较低,处于较好的保种状态,具有较大的品种资源开发潜力。     

香樟ISSR-PCR反应体系的正交优化

应用L9(34)正交表建立了适合于樟树ISSR-PCR的优化反应体系,即20μL ISSR反应体系中含有1×PCR buffer、dNTP0.25 mmol/L、引物0.3μmol/L、Mg2 2.0 mmol/L、Taq酶1 U和模板DNA50 ng.适宜的扩增条件为:94℃预变性4 min;94℃变性40 s,58℃退火1 min,72℃延伸2 min,42个循环;72℃延伸10 min;4℃保存.     

DNA分子标记技术在食用菌研究中的应用及进展

根据近年来国内外的文献报道,对DNA分子标记技术在食用菌遗传育种、菌种和菌株鉴定、遗传多样性分析、基因定位和克隆等研究中的应用及进展进行了综述和分析.认为随着分子生物学技术的进一步发展,将有更多、更有效的分子标记在食用菌研究中得以应用,同时随着分子标记技术应用的深入,更高密度、更为实用有效的遗传连锁图谱将被建立,大量重要的功能基因将被定位和克隆,这将为进一步培育高产、优质、抗病、耐贮等食用菌新品种奠定良好的基础.     

紫色甘薯资源的形态学和ISSR荧光标记分析

【目的】分析紫色甘薯品种的遗传多样性,其遗传差异,为紫色甘薯遗传育种亲本选择提供理论依据。【方法】以引进的10份紫色甘薯为材料,利用形态学性状和ISSR 荧光标记毛细管电泳技术分析品种遗传多样性。【结果】基于20项形态学性状的聚类图,在欧式距离为9.97处,可分为两大类群,第一大类为宁紫1号、山东紫薯、渝紫2号、徐紫8号、越南紫薯 ;第二大类为鄂12、宁紫4号、渝紫3号、渝紫11、南紫018 。紫色甘薯种质资源的观测等位基因为(Na)为2个,平均有效等位基因数(Ne)为1.919 0,期望杂合度(He)为0.237 5,香农指数(I)为0.433 3,在遗传距离为0.96时,10份供试紫色甘薯材料可分为两大类。【结论】部分供试材料的ISSR标记聚类结果与形态学标记在遗传背景和类群划分上具有一致性,但是两种鉴定分类方法的聚类分析结果也存在一定差异。紫色甘薯种质材料遗传多样性丰富,将形态学及ISSR标记结合能有效提高品种间特异性的鉴定,更能客观、准确的确定种质资源的亲缘关系。